Sortase-mediated modification of alpha DEC205 affords optimization of antigen presentation and immunization against a set of viral epitopes

作者：Swee Lee Kim; Guimaraes Carla P; Sehrawat Sharvan; Spooner Eric; Barrasa M Inmaculada; Ploegh Hidde L^*

来源：Proceedings of the National Academy of Sciences of the United States of America, 2013, 110(4): 1428-1433.

DOI：10.1073/pnas.1214994110

• antibody conjugates
• vaccines
• vaccine development
• protein engineering

A monoclonal antibody against the C-type lectin DEC205 (alpha DEC205) is an effective vehicle for delivery of antigens to dendritic cells through creation of covalent alpha DEC205-antigen adducts. These adducts can induce antigen-specific T-cell immune responses or tolerance. We exploit the transpeptidase activity of sortase to install modified peptides and protein-sized antigens onto the heavy chain of alpha DEC205, including linkers that contain nonnatural amino acids. We demonstrate stoichiometric site-specific labeling on a scale not easily achievable by genetic fusions (49 distinct fusions in this report). We conjugated a biotinylated version of a class I MHC-restricted epitope to unlabeled alpha DEC205 and monitored epitope generation upon binding of the adduct to dendritic cells. Our results show transfer of alpha DEC205 heavy chain to the cytoplasm, followed by proteasomal degradation. Introduction of a labile dipeptide linker at the N terminus of a T-cell epitope improves proteasome-dependent class I MHC-restricted peptide cross-presentation when delivered by alpha DEC205 in vitro and in vivo. We also conjugated alpha DEC205 with a linker-optimized peptide library of known CD8 T-cell epitopes from the mouse gamma-herpes virus 68. Animals immunized with such conjugates displayed a 10-fold reduction in viral load.

• 出版日期2013-1-22